Title: IGF-I as a mediator of skeletal anabolic response to loading It is well known that the development of skeletal architecture and maintenance of bone mass are dependent on mechanical stimulation. Although gene expression studies have revealed evidence for the involvement of several potential candidate genes as mediators of mechanical loading (ML) response, direct evidence for the involvement of these genes in mediating bone anabolic response to ML is lacking. In terms of potential mediators of ML response in the skeleton, Insulin like growth factor-I (IGF-I) has received considerable attention since IGF-I expression has been shown to increase rapidly in response to mechanical strain in bones in vivo and bone cells in vitro and since transgenic studies involving disruption of IGF-I gene have shown that IGF-I is a key bone formation regulator. Based on past findings and our preliminary data, we propose the hypothesis that ML induced increase in IGF-I expression interacts with estrogen to determine the amount of new bone formed in response to mechanical strain. To evaluate this hypothesis, we have proposed cause and effect relationship studies between IGF-I, estrogen and bone formation response to ML utilizing IGF-I conditional knockout (KO) mice in which IGF-I gene is disrupted using Cre/LoxP approach. We will generate IGF-I KO mice by crossing loxP IGF-I mice with transgenic Cre mice in which Cre expression is driven by promoter and enhancer elements of alpha2 chain of type I collagen promoter. ML will be induced by four-point bending in the tibia or axial compression of forelimb. In vivo pQCT and Real time RT-PCR will be used to quantitate changes in bone parameters (loaded and non-loaded bones) and gene expression, respectively. Three-point bending and micro-CT will be used to measure changes in the mechanical properties, and architecture in response to loading in both the KO and the control mice. Histology will be carried out using calcein-double labeling in order to measure bone formation rate, osteoblast cell population, mineral apposition rate, and osteoblast activity resulting from ML. To determine if estrogen interacts with IGF-I to regulate skeletal anabolic response to loading, control and IGF-I conditional KO mice will be ovariectomized and skeletal response to ML determined. Our understanding of the role of IGF-I and estrogen in mediating bone formation response to loading will lead to development of strategies to increase bone anabolic response to ML by modulating local IGF-I actions in bone.